Towards non-invasive imaging of vulnerable atherosclerotic plaques by targeting co-stimulatory molecules.

BACKGROUND: Myocardial infarction and stroke are the life-threatening consequences after plaque rupture in coronary or carotid arteries. Positron emission tomography employing [(18)F]fluorodeoxyglucose can visualize plaque inflammation; however, the question remains whether this is specific for plaque vulnerability. The pathophysiology of vulnerable plaques suggests several molecular processes. Here, we propose the co-stimulatory molecules CD80 and CD86 as potential new targets for non-invasive imaging. METHODS AND RESULTS: Human atherosclerotic segments were obtained from carotid endarterectomy and classified into stable and vulnerable plaques. We identified CD80 and CD86 with significantly higher mRNA levels in vulnerable than stable plaques. CD80+ and CD86+ cells were found in spatial proximity to CD83+ dendritic cells and CD68+ macrophages of atherosclerotic plaques. As a proof of target-expression we labeled a low molecular weight ligand, which has a high affinity for human CD80, with carbon-11 to perform in vitro autoradiography with human plaque slices. We observed 3-fold higher binding to vulnerable than stable plaques, demonstrating a first approach towards discriminating between the two plaque types. Positron emission tomography studies showed accumulation in CD80+ Raji xenografts, low radioactivity in myocardium and rapid clearance from the blood pool in mice. CONCLUSION: In human carotid arteries, the co-stimulatory molecules CD80 and CD86 show significantly higher expression levels in vulnerable compared to stable plaques. With the novel CD80-specific radiotracer we are able to discriminate between stable and vulnerable atherosclerotic plaques in vitro. This is an important step towards non-invasive imaging of the life-threatening vulnerable lesions in humans.

Resolution, sensitivity and precision with autoradiography and small animal positron emission tomography: implications for functional brain imaging in animal research.

Quantitative autoradiographic methods for in vivo measurement of regional rates of cerebral blood flow, glucose metabolism, and protein synthesis contribute significantly to our understanding of phsysiological and biochemical responses of the brain to changes in the environment. A disadvantage of these autoradiographic methods is that experimental animals can be studied only once. With the advent of small animal positron emission tomography (PET) and with increases in the sensitivity and spatial resolution of scanners it is now possible to use adaptations of these methods in experimental animals with PET. These developments allow repeated studies of the same animal, including studies of the same animal under different conditions, and longitudinal studies. In this review we summarize the tradeoffs between the use of autoradiography and small animal PET for functional brain imaging studies in animal research.

Radioautographology general and special.

A new concept, termed "radioautographology" is advocated and its contents are reviewed. This term is the coinage synthesized from "radioautography" and "(o)logy", expressing a new science derived from radioautography. The concept of radioautographology (RAGology) is a science to localize the radioactive substances in the biological structure of the objects and to analyze and to study the significance of these substances in the biological structure. On the other hand, the old term radioautography (RAG) or autoradiography (ARG) is the technique to demonstrate the pattern of localization of various radiolabeled compounds in biological specimens. The specimens used in biology and medicine are cells and tissues. They are fixed, sectioned and made contact with the radioautographic emulsions, exposed and developed to produce metallic silver grains. Such specimens are designated as radioautographs (or autoradiographs) and the patterns of pictures made of silver grains are named radioautograms. Those people who produced radioautographs were formerly named radioautographers (or autoradiographers) who were only technicians, while those who study RAGology are not technicians but scientists and should be called as radioautographologists. The science of radioautographology was developed in the 20th century and can be divided into two parts, general radioautographology and special radioautographology, as most natural sciences usually can. The general radioautographology is the technology of RAG which consists of 3 fields of sciences, physics concerning radioactivity, histochemistry treating the cells and tissues and photochemistry dealing with the photographic emulsions. The special radioautographology, on the other hand, consists of applications of general radioautographology to various biological and medical sciences. The applications can be classified into several scientific fields, i.e., cellular molecular biology, anatomy, histology, embryology, pathology and pharmacology. Studies carried out in our laboratory were summarized and reviewed. The results obtained from the technology includes 4-dimensional structures of the organs taking the time dimension into account by labeling cells and localizing the sites of incorporation, synthesis, discharge of the labeled compounds in connection with the time lapse and aging of animals. All the results obtained from such applications should be systematized as a new filed of science in the future in the 21st century.

Receptor binding techniques.

This overview first discusses issues relating to the selection of radioligand for receptor binding assays, including the isotopic label and considerations pertaining to the pharmacological and chemical profile of the ligand. This is followed by a section on characterization of ligand-binding assays, starting with tissue preparation methods, followed by detection of specific binding, determination of incubation and washing conditions and a discussion of saturation and competition assay formats. Quantification of the assay results can be accomplished by autoradiography or film densitometry. Finally, methods and considerations for analysis of the resulting data are presented.

Metabolic mapping.

This unit describes a high-resolution [(3)H]2-deoxyglucose (2DG)/immunohistochemistry double labeling protocol which enables simultaneous visualization of metabolic and immunohistochemical markers (e.g., neurotransmitter-specific epitopes) in the same tissue section. This approach generates single-cell metabolic measures from large (many thousands) samples of neurons, and allows the assignment of putative neurotransmitter types to large samples of functionally assayed single neurons. Thus, it provides a unique opportunity to assay the metabolic activities of histochemically identified neural elements throughout an entire pathway. [(3)H]2DG injection and perfusion fixation of hamsters or mice is described first, followed by immunohistochemical staining of brain slices for a monoclonal antibody against glutamic acid decarboxylase-6 (GAD-6). Included are the steps for dipping and developing slides for autoradiography. An alternate protocol gives details of staining for cytochrome oxidase (CO).